Hematopoietic Deficiency of miR-223 Attenuates Thrombosis in Response to Photochemical Injury in Mice

Hui Wang,Chiao Guo,Daniel T Eitzman,Kyle Kleiman,Qian Wang

doi:10.1038/s41598-017-01887-x

Abstract

Some studies have shown that levels of MicroRNA (miR)-223 derived from platelets in the plasma are reduced following inhibition of platelet function, while others have shown a correlation between low plasma miR-223 and high on-treatment platelet reactivity. The present study seeks to investigate the role of miR-223 in arterial thrombosis. A model of photochemical-induced carotid thrombosis was applied to miR-223 deficient mice and littermate (WT) controls. Mice deficient in miR-223 exhibited significantly prolonged times to occlusive thrombosis compared to WT mice indicating a protective effect of miR-223 deficiency. Bone marrow transplantation experiments confirmed that the hematopoietic pool of miR-223 was responsible for differences in thrombosis times. Transfusion of either WT platelets or extracellular vesicles derived from WT platelets were both sufficient to shorten thrombosis times in miR-223 deficient recipients. The effect of platelet transfusions on IGF-1R was explored. These experiments revealed that vascular IGF-1R was down-regulated by platelet miR-223. Furthermore, inhibition of IGF-1R abolished the protection conferred by miR-223 deficiency on thrombosis. In conclusion, platelet miR-223 is a regulator of arterial thrombosis following endothelial injury through effects on vascular wall IGF-1R. This study indicates that platelet miR-223 is a potential therapeutic target for prevention of arterial thrombosis.

Highlights

Arterial thrombosis was defined as flow cessation for at least 10 minutes
To determine whether extracellular vesicles (ECVs) isolated from platelets were sufficient to affect the thrombosis phenotype, we examined the effect of ECV transfusion, isolated from WT or miR-223 deficient platelets, to recipient miR-223 deficient mice on arterial thrombosis
Consistent with previous studies that have demonstrated hematopoietic miR-223 may be transferred to other cell types such as endothelial cells[8, 9], and affect properties of endothelial cells[14], we demonstrated that platelet-to-endothelial transfer of miR-223 regulates thrombosis and that this effect is related to changes in vascular insulin growth factor-1 receptor (IGF-1R) expression

Summary

Results and Discussion

To investigate the role of miR-223 on arterial thrombosis following endothelial injury, we performed rose bengal photochemical injury to the carotid arteries of WT and miR-223 deficient mice. MiR-223 deficient mice exhibited significantly prolonged times to occlusive thrombosis compared to WT mice (Fig. 1A), indicating a protective effect of miR- 223 deficiency on thrombosis in this model. The time to occlusive thrombosis in WT mice receiving bone marrow from miR-223 deficient mice was markedly prolonged and similar to times observed in non-transplanted miR223 deficient mice (Fig. 2) These data establish the hematopoietic pool of miR-223 as playing a regulatory role in carotid thrombosis following endothelial injury. After washing bEnd[3] cells and isolating RNA, expression of miR-223 was increased in bEnd[3] cells incubated with WT ECVs (Fig. 4A) To validate this platelet-to-endothelial miR-223 transfer in a more physiological setting, miR-223 expression was analyzed in the carotid arterial wall following injury from miR-223 deficient recipients that received WT platelet transfusions. This study indicates that inhibition of platelet miR-223, or activation of its downstream target IGFR1, are potential therapeutic strategies for prevention of arterial thrombosis

Methods

Author Contributions

Additional Information